Since the inadvertent exposure of large numbers of people to the simian polyomavirus SV40 in the late 1950s and early 1960s, there has been concern of a possible oncogenic role for this virus in brain tumors as well as other human malignancies. Several studies have proven the oncogenic potential of polyomaviruses in nonhost experimental animals; however, studies concerning the association between human brain tumors and simian and human polyomaviruses have yielded inconclusive results. We have investigated the relationship of SV40 to a malignant astrocytoma found in the right frontal lobe of a pigtail macaque (Macaca nemestrina) infected with simian immunodeficiency virus (SIV). Consistent with the histologic diagnosis, the tumor was immunoreactive with antibodies to S-100 protein, vimentin, and glial fibrillary acidic protein, but negative for neurofilament protein, synaptophysin, neuron-specific enolase, and chromogranin A. At the time of SIV inoculation the animal was seropositive for SV40. Polymerase chain reaction (PCR) assay of tumor DNA, but not normal brain DNA, yielded a 300 base-pair fragment corresponding to